Prediction of antibody structural epitopes via random peptide library screening and next generation sequencing

Antibodies are essential to functional immunity, yet the epitopes targeted by antibody repertoires remain largely uncharacterized. To aid in characterization, we developed a generalizable strategy to predict antibody-binding epitopes within individual proteins and entire proteomes. Specifically, we selected antibody-binding peptides for 273 distinct sera out of a random library and identified the peptides using next-generation sequencing. To predict antibody-binding epitopes and the antigens from which these epitopes were derived, we tiled the sequences of candidate antigens into short overlapping subsequences of length k (k-mers). We used the enrichment over background of these k-mers in the antibody-binding peptide dataset to predict antibody-binding epitopes. As a positive control, we used this approach, termed K-mer Tiling of Protein Epitopes (K-TOPE), to predict epitopes targeted by monoclonal and polyclonal antibodies of well-characterized specificity, accurately recovering their known epitopes. K-TOPE characterized a commonly targeted antigen from Rhinovirus A , predicting four epitopes recognized by antibodies present in 87% of sera (n = 250). An analysis of 2,908 proteins from 400 viral taxa that infect humans predicted seven enterovirus epitopes and five Epstein-Barr virus epitopes recognized by >30% of specimens. Analysis of Staphylococcus and Streptococcus proteomes similarly predicted 22 epitopes recognized by >30% of specimens. Twelve of these common viral and bacterial epitopes agreed with previously mapped epitopes with p-values < 0.05. Additionally, we predicted 30 HSV2-specific epitopes that were 100% specific against HSV1 in novel and previously reported antigens. Experimentally validating these candidate epitopes could help identify diagnostic biomarkers, vaccine components, and therapeutic targets. The K-TOPE approach thus provides a powerful new tool to elucidate the organisms, antigens, and epitopes targeted by human antibody repertoires.

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“…The bacterial peptide display screening protocol was carried out as previously described [29,67]. Briefly, an E .…”

Section: Methodsmentioning

confidence: 99%

A general approach for predicting protein epitopes targeted by antibody repertoires using whole proteomes

et al. 2019

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“…The bacterial peptide display screening protocol was carried out as previously described [29,62]. Briefly, an E. coli library displaying approximately 8 billion different 12-mer peptides was combined with 1:100 diluted serum.…”

Section: Methodsmentioning

confidence: 99%

A general approach for identifying protein epitopes targeted by antibody repertoires using whole proteomes

Paull

Johnston

Ibsen

et al. 2019

Preprint

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28Antibodies are essential to functional immunity, yet the epitopes targeted by antibody 29 repertoires remain largely uncharacterized. To aid in characterization, we developed a 30 generalizable strategy to identify antibody-binding epitopes within individual proteins and entire 31 proteomes. Specifically, we selected antibody-binding peptides for 273 distinct sera out of a 32 random library and identified the peptides using next-generation sequencing. To identify 33 antibody-binding epitopes and the antigens from which these epitopes were derived, we tiled the 34 sequences of candidate antigens into short overlapping subsequences of length k (k-mers). We 35 used the enrichment over background of these k-mers in the antibody-binding peptide dataset to 36 identify antibody-binding epitopes. As a positive control, we used this approach, termed K-mer 37 Tiling of Protein Epitopes (K-TOPE), to identify epitopes targeted by monoclonal and polyclonal 38 antibodies of well-characterized specificity, accurately recovering their known epitopes. K-39 TOPE characterized a commonly targeted antigen from Rhinovirus A, identifying three epitopes 40 recognized by antibodies present in 83% of sera (n = 250). An analysis of 2,908 proteins from 41 400 viral taxa that infect humans revealed seven enterovirus epitopes and five Epstein-Barr virus 42 epitopes recognized by >30% of specimens. Analysis of Staphylococcus and Streptococcus 43 proteomes similarly revealed six epitopes recognized by >40% of specimens. These common 44 viral and bacterial epitopes exhibited excellent agreement with previously mapped epitopes.45 Additionally, we identified 30 HSV2-specific epitopes that were 100% specific against HSV1 in 46 novel and previously reported antigens. The K-TOPE approach thus provides a powerful new 47 tool to elucidate the organisms, antigens, and epitopes targeted by human antibody repertoires. 48 49 50 Introduction 51 52 Immunological memory allows for rapid antibody responses towards diverse antigens 53 long after initial exposure. For example, the adaptive immune response to many vaccinations is 54 often sustained throughout an individual's lifetime [1]. This immunological information is 55 archived within the genes encoding B-cell and T-cell receptors along with the corresponding 56 receptor structures, but has proven difficult to characterize in a comprehensive manner. The 57 ability to more fully interrogate immunological memory could reveal exposures to pathogens, 58 commensal organisms, and allergens. Such information has proven useful for correlating 59 antibody responses with disease outcomes to design more effective vaccines [2]. A detailed 60 record of immune exposures can also facilitate the identification of biomarkers to diagnose 61 infectious [3], autoimmune [4], and allergic conditions [5]. Furthermore, the capability to 62 broadly characterize antibody repertoires at the epitope level could be used to identify conserved 63 pathogen epitopes [6] and tumor specific antigen epitopes [7] to aid in therapeutic disco...

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“…Indeed, the application of CAR-based therapy has not been investigated thoroughly in autoimmune diseases because there is no appropriate antigens to target them on the aberrant autoreactive cells selectively. By the advent of next-generation sequencing (NGS), different types of epitopes can be predicted and identified for future CAR-based immunotherapy (123). In addition, genome editing technology, CRISPR/Cas9, has a great influence on detecting hidden self-antigens in autoimmune diseases to increase the specificity of CAR-T cells (124).…”

Section: Car-based Therapies: Future Perspectivementioning

confidence: 99%

Chimeric Antigen Receptor Based Therapy as a Potential Approach in Autoimmune Diseases: How Close Are We to the Treatment?

et al. 2020

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Despite significant breakthroughs in understanding of immunological and physiological features of autoimmune diseases, there is currently no specific therapeutic option with prolonged remission. Cell-based therapy using engineered-T cells has attracted tremendous attention as a practical treatment for autoimmune diseases. Genetically modified-T cells armed with chimeric antigen receptors (CARs) attack autoreactive immune cells such as B cells or antibody-secreting plasma cells. CARs can further guide the effector and regulatory T cells (Tregs) to the autoimmune milieu to traffic, proliferate, and exert suppressive functions. The genetically modified-T cells with artificial receptors are a promising option to suppress autoimmune manifestation and autoinflammatory events. Interestingly, CAR-T cells are modified to a new chimeric auto-antibody receptor T (CAAR-T) cell. This cell, with its specific-antigen, recognizes and binds to the target autoantibodies expressing autoreactive cells and, subsequently, destroy them. Preclinical studies of CAR-T cells demonstrated satisfactory outcomes against autoimmune diseases. However, the lack of target autoantigens remains one of the pivotal problems in the field of CAR-T cells. CAR-based therapy has to pass several hurdles, including stability, durability, trafficking, safety, effectiveness, manufacturing, and persistence, to enter clinical use. The primary goal of this review was to shed light on CAR-T immunotherapy, CAAR-T cell therapy, and CAR-Treg cell therapy in patients with immune system diseases.

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Prediction of antibody structural epitopes via random peptide library screening and next generation sequencing

Cited by 15 publications

References 30 publications

A general approach for predicting protein epitopes targeted by antibody repertoires using whole proteomes

A general approach for predicting protein epitopes targeted by antibody repertoires using whole proteomes

A general approach for identifying protein epitopes targeted by antibody repertoires using whole proteomes

Chimeric Antigen Receptor Based Therapy as a Potential Approach in Autoimmune Diseases: How Close Are We to the Treatment?

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